Some significant differences in wall chemistry among four commercial Agaricus bisporus strains

Significant differences in gross wall chemical composition were detected in four commercial Agaricus bisporus strains. All were grown under the same conditions and their walls prepared by a mild method of breakage. A more detailed analysis of the wall fractions, isolated by means of their distinct solubilities, also showed striking structural differences among the four strains studied. The detected differences, not only in the overall composition of the wall but also in the polysaccharide structure, could assist in the characterization of strains and/or varieties of the commercial basidiomycete A. bisporus.     

Chemical Analysis of the Lamella Walls of Agaricus bisporus Fruit Bodies

Purified lamella wall fragments of Agaricus bisporus fruit bodies were analyzed and shown to consist of neutral sugars (46.5%), hexosamines (31.7%), proteins (9.5%), some lipid material (10.0%), and ash (1.4%). The cell walls were fractionated on the basis of their polysaccharide solubility in water and alkaline solutions. The isolated fractions, using methylation analysis, exhibited striking chemical structural differences compared with the same fractions obtained from the corresponding vegetative cells and fruit bodies (stipe and pileus) walls. The structural differences detected in the wall seem to correspond to the ultimate differentiation of the mycelium inside the fruit body of A. bisporus. Received: 30 November 1998 / Accepted: 29 January 1999     

Comparative compositional analysis of walls with two different morphologies: archetypical versus transfer-cell-like

Summary Abnormally thick cell walls of a clonal maize cell line with the labyrinth wall morphology found in transfer cells were analyzed and compared to the relatively thin and even archetypical walls of a sister cell line. Despite a drastic difference in wall morphology between the transfer and archetypical cell walls, the chemical composition of the walls was essentially the same. There were no major differences in the glycosyl residue composition, in the amount of total lipid, and in the amount of total protein. The amounts of wall material released by chemical extraction of cellulosic, hemicellulosic, and pectic fractions were the same for the two types of walls. There were some differences in the protein profile and in the inorganic ion content between the transfer and archetypical walls. These results indicate that profound changes in wall morphology can be brought about in the absence of gross changes in wall composition and suggest that major changes in time- or place-dependent deposition and/or subtle changes in arrangement of rare wall constituents may be responsible.     

STRUCTURE AND COMPOSITION OF THE RESISTANT SPORANGIAL WALL IN THE FUNGUS ALLOMYCES

The fine structure and chemical composition of the wall of resistant sporangia of Allomyces neo-moniliformis were investigated. Studies with the electron microscope showed that the wall is approximately 1.3 μ in thickness and is of complex construction. It consists essentially of three parts: a five-layered outer wall, two layers of “cementing substances,” and a single-layered inner wall. The presence of a highly convoluted cell membrane was also demonstrated. Six structural components were found to make up the walls of the resistant sporangia: glucose, glucosamine, chitin, melanin, protein, and lipids. Comparison of the structure and composition of the walls of resistant sporangia with the walls of hyphae and zoospores of Allomyces as reported by other investigators showed that, while the structure is very different, the composition is quite similar with only melanin and lipids apparently being absent from the zoospore and hyphal walls.     

Cell wall composition of the yeast and mycelial forms of Yarrowia lipolytica

The cell walls of the yeast and mycelial forms of Yarrowia lipolytica were isolated and purified. Electron microscopy studies showed no differences between both types of cell walls. Chemical analysis revealed that the yeast cell wall contained 70% neutral carbohydrate, 7% amino sugars, 15% protein, 5% lipids and 0.8% phosphorus. Mycelial cell walls contained 70% carbohydrate, 14% aminosugars, 6% protein, 5% lipids and 0.6% phosphorus. Three polysaccharides: -glucan, mannan and chitin were detected. Proteins were solubilized from both cell wall fractions and separated by polyacrylamide gel electrophoresis. About 50 protein bands were detected, four of them corresponding to glycoproteins. The cell walls of the yeast and mycelial forms of Y. lipolytica were qualitatively similar and only quantitative differences were found.Abbreviations GlcNAc N-acetylglucosamine - FITC-WGA fluorescein isothiocyanate-wheat germ agglutinin - PAS periodic acid Schiff     

Differentiation and wall chemistry of Agaricus bisporus vegetative and aggregated micelia

Changes in the chemical composition of isolated cell walls and fractions were encountered during the differentiation of vegetative and aggregated mycelia of Agaricus bisporus.Differentiation was accompanied by quantitative variations of the wall polysaccharidic components. Neutral carbohydrates were composed of glucose, galactose, mannose and xylose and glucosamine as the only amino sugar. Differences in wall chemistry were correlated to the secondary and tertiary mycelial forms.     

Comparative studies onChlorella cell walls: Induction of protoplast formation

Among 12 strains ofChlorella ellipsoidea, C. vulgaris, andC. saccharophila tested, 4 strains (1,C. ellpsoidea; 2,C. vulgaris; 1,C. saccharophila) formed osmotically labile protoplasts after treatment with mixtures of polysaccharide degrading enzymes. The relationship between enzymatical digestibility and structure or composition ofChlorella cell walls were studied by electron microscopy and staining techniques with some specific dyes. The cell wall structures of the 12Chlorella strains were grouped into three types: (1) with a trilaminar outer layer, (2) with a thin outer monolayer, and (3) without an outer layer. Protoplasts were formed only from the strains with a cell wall of Type 2. In the strains with a cell wall of Type 1, the outer layer protected the inner major microfibrillar layer against enzymatic digestion. The cell wall of Type 3 was totally resistant to the enzymes; the chemical composition of the cell wall would be somewhat different from that of other types.     

Studies on the Cell Wall of Chlorella V. Comparison of the Cell Wall Chemical Compositions in Strains of Chlorella ellipsoidea

Cell walls of four strains of Chlorella ellipsoidea (IAM C-27,C-87, C-102 and C-183) were compared as to their chemical compositions.Many differences were found: (1) The sugar composition of alkali-soluble cell walls differedin quantity as well as quality with glucuronic acid being foundonly in C-27 and C-87. (2) In alkali-insoluble cell walls glucosamine was found onlyin C-27. The other three strains contained mainly glucose. (3) The amino acid compositions of the alkali-insoluble cellwalls markedly differed among the four strains. The cell wallof C-102 contained more amino acids than carbohydrates, butC-27 and C-87 contained extremely little amino acid. In addition to the variation in cell wall composition, the opticalanisotropy findings also differed for these cell walls of Chlorellastrains which had been grouped as the same species. (Received August 16, 1983; Accepted December 27, 1983)     

Study of supramolecular structures released from the cell wall of Candida albicans by ethylenediamine treatment

Candida albicans cell wall components were analyzed by ethylenediamine (EDA) treatment. Based on their different solubility properties, the cell wall components produced three fractions (A, B, and C). Fractions B (EDA-soluble, water-insoluble) and C (EDA-insoluble) contained glucan, chitin, and protein in different proportions. After zymolyase (mainly a β-glucanase complex) or chitinase treatment of fractions B and C, more polysaccharides and proteins were solubilized by a second EDA treatment, suggesting that the solubility of the polymers in EDA depends on the degree of polymer interactions. Western blot analysis using two monoclonal antibodies (1B12 and 4C12) revealed electrophoretic patterns that were similar in mycelial and yeast morphologies, except that in material obtained from mycelial walls, an additional band was detected with MAb 1B12. Fluorescence microscopy of cell wall fractions treated with FITC-labeled Con-A, Calcofluor white, and FITC-labeled agglutinin showed that glucan and mannoproteins are uniformly distributed in fractions B and C, while chitin is restricted to distinct patches. Transmission electron microscopy demonstrated that fraction C maintained the original shape of the cells, with an irregular thickness generally wider than the walls. When fraction C was treated with chitinase, the morphology was still present and was maintained by an external glucan layer, with an internal expanded fibrillar material covering the entire cellular lumen. Degradation of the glucan skeleton of fraction C with zymolyase resulted in the loss of the morphology. Received: 1 April 1996 / Accepted: 2 September 1996     

Comparative investigation of primary and tertiary endodermal cell walls isolated from the roots of five monocotyledoneous species: chemical composition in relation to fine structure

The chemical composition of isolated endodermal cell walls from the roots of the five monocotyledoneous species Monstera deliciosa Liebm., Iris germanica L., Allium cepa L., Aspidistra elatior Bl. and Agapanthus africanus (L.) Hoffmgg. was determined. Endodermal cell walls isolated from aerial roots of M. deliciosa were in their primary developmental state (Casparian bands). They contained large amounts of lignin (6.5% w/w) and only traces of suberin (0.5% w/w). Endodermal cell walls isolated from the other four species were in their tertiary developmental state. Lignin was still the more abundant cell wall polymer with amounts ranging from 3.8% (w/w, A. cepa) to 4.5% (w/w, I. germanica). However, compared to endodermal cell walls in their primary state of development (Casparian bands), tertiary endodermal cell walls contained significantly higher amounts of suberin, ranging from 1.8% (w/w, I. germanica) to 3.0% (w/w, A. africanus). Thus, chemical characterization of endodermal cell walls from five different species revealed that lignin was the dominant cell wall polymer in the Casparian band of M. deliciosa, whereas tertiary endodermal cell walls contained, in addition to lignin, increasing amounts of suberin (I. germanica, A. cepa, A. elatior and A. africanus). Besides the two biopolymers lignin and suberin, cell wall carbohydrates in the range of between 40 and 60% were also quantified. The sum of all cell wall compounds investigated by gas chromatography resulted in a recovery of 50–80% of the dry weight of the isolated cell wall material. Quantitative chromatographic results in combination with microscopic studies are consistent with the existence of a distinct suberin lamella and lignified tertiary wall deposits. From these data it can be concluded that the barrier properties of the endodermis towards the apoplastic transport of ions and water will increase from primary to tertiary endodermal cell walls due to their increasing amounts of suberin. Received: 23 August 1997 / Accepted: 28 January 1998     

Comparative Chemical Characterization of Pigmented and Less Pigmented Cell Walls of Alternaria tenuissima

Alternaria tenuissima, the parasitic fungus, was obtained from the pruned upper-cut surfaces of mulberry stems. This fungus contains dark pigment because of the presence of melanin in the cell wall. To obtain less-pigmented cell walls, this fungus was grown under dark condition. When the pigmented and less-pigmented cell walls were chemically analyzed, no differences were observed in amino-acid composition, hexoses, or pentoses. However, in pigmented cell walls, higher contents of melanin (2.6%) were found than in less-pigmented cell walls (0.3%). Interestingly, a significant difference was observed in the relative fatty-acid compositions between these two types of cell walls. Among the major fatty acids, there were increased concentrations of tetradecanoic acid (C14:0), hexadecanoic acid (C16:0), 9-hexadecenoic acid (C16: 1,Δ9), and 9-octadecanoic acid (C18:1,Δ9) and a concomitant decrease in 9,12-octadecadienoic acid (C18:2,Δ9,12) in less-pigmented compared with pigmented cell walls. This difference in fatty-acid composition may be related to the higher percentage of melanin in the pigmented than the less-pigmented cell walls. Lesser amounts of 9,12-octadecadienoic acid in less-pigmented cell walls may have been caused by the growth of the fungus under environmental stress conditions. An interesting observation was the presence in pigmented cell walls only of methyl-substituted fatty acids with carbon numbers C14 to C17, but their occurrence could not be ascertained in the present study.     

Natural Abundance 13C Nuclear Magnetic Resonance Spectra of Intact Fungal Mycelia

We examined the change of the composition of the cell wall polysaccharides prepared from cells of the salt-tolerant yeast Zygosaccharomyces rouxii grown in two media containing 20% NaCl and 0% NaCl. Comparative analysis of their walls showed that the wall obtained from salt-free medium had greater quantities of alkali-insoluble fraction and smaller quantities of mannan than the walls obtained from 20% NaCl medium. The alkali-insoluble fractions from the cell walls obtained from salt-free medium contained a large amount of glucosamine and a smaller amount of linear β-1,3-glucosidic linkage than the fractions from the cell walls obtained from 20% NaCl medium. Structural analyses showed that the mannans from each cell wall contained an α-1,6-mannbsidic linked backbone to which single mannose and mannobiose units were connected as side chains by α-1,2-mannosidic linkages. However, when cells were grown in the presence of 20% NaCl, the side chains of the mannans consisting of a mannobiose unit were largely reduced.

These results indicated that the structure of alkali-insoluble glucan and mannan were greatly affected by the presence of NaCl in the final medium.     

Cell wall composition and synthesis via Golgi-directed scale formation in the marine eucaryote,Schizochytrium aggregatum,with a note on Thraustochytrium sp.

Summary Cell walls of Schizochytrium aggregatum and Thraustochytrium sp. were mechanically isolated and subjected to chemical analysis. On a dry weight basis the cell walls contain 21–36% carbohydrate and 30–43% protein. The principal sugar (>95%) of the Schizochytrium wall is l-galactose, while the Thraustochytrium cell wall contains l-galactose, d-galactose and xylose with l-galactose predominating. Ultrastructurally the cell walls of both organisms consist of a laminated structure which yields thin, flexible, nearly circular scales (0.5–1.1 in diameter) upon sonic disintegration. Structures presumed to be developing wall scales are found within cisternae of the Golgi apparatus in both organisms. The chemical composition and method of formation of the cell wall in these two protists is distinctly different from that found in the Saprolegniales (Oomycetes), the group with which these organisms have hitherto been aligned.     

Comparison of the surface structures of choline-less Torulopsis pintolopesii grown on defined media with choline or methionine

Summary The chemical composition of cell walls from choline-less Torulopsis pintolopesii grown with choline or with methionine was studied. Methioninegrown cells synthesized a weakened cell wall compared to normal choline-grown yeast. The ethylenediamine fractionation procedure yielded three fractions—A, B, and C—with different solubilities. Glucose and mannose were detected in hydrolysed unfractionated cell walls from yeasts grown under both conditions as well as in all fractions. Glucose content was greater in fractions B and C from methioninegrown cells; the mannose content was about the same. Walls from choline-grown cells (W _c ) had 25% more protein than walls from methionine-grown cells (W _m ). The amino acid composition of the proteins of W _c and W _m was not qualitatively altered. Seventeen amino acids were identified; glutamic and aspartic acids and valine predominated. W _c had 3.5 times more lipid than W _m . The amount of phosphorus was the same. Yeasts grown on methionine synthesized more ergosterol than choline-grown cells. The rate of formation of spheroplasts was higher in methionine-grown cells. Rates of incorporation of adenine, glutamic acid, and uracil were similar in cells grown on methionine or choline; incorporation of phenylalanine and tyrosine was depressed in methionine-grown cells.     

Hyphal walls of isolated lichen fungi

The hyphal walls of three mycobionts, isolated from the lichens Xanthoria parietina, Tornabenia intricata and Sarcogyne sp. were investigated by two techniques: microautoradiography of fungal colonies exposed to radioactive carbohydrate precursors; and binding, in vivo, of fluorescein conjugated lectins to hyphal walls of such colonies.N-[³H] acetylglucosamine was readily incorporated into tips, young hyphal walls and septa of the three mycobionts and the free-living fungus Trichoderma viride, but not into Phytophthora citrophthora, indicating that chitin is a major component of the mycobionts' hyphal walls. All three mycobionts, but neither of the free-living fungi, incorporated [³H] mannose and [³H] mannitol into their hyphal walls.Fluorescein-conjugated wheat germ agglutinin was bound to the hyphal walls of the three mycobionts and T. viride, but not to the walls of P. citrophthora; the binding pattern was similar to the grain pattern obtained in autoradiographs after short N-[³H] acetylglucosamine labelling. As wheat germ agglutinin binds specifically to chitin oligomers, the lectin binding tests further confirmed that chitin is a mycobiont hyphal wall component.Binding characteristics of several fluorescein-conjugated lectins to the three mycobionts indicated that this technique can yield useful information concerning the chemical composition of hyphal wall surfaces.List of abbreviations FITC fluorescein isothiocyanate - WGA wheat germ agglutinin - TCA trichloroacetic acid - PNA peanut agglutinin - LA lotus agglutinin - Glc NAc N-acetylglucosamine - ConA concanavalin A - SBA soybean agglutinin - WBA waxbean agglutinin Part of an M.Sc. thesis submitted by A. Braun to the Department of Botany, Tel Aviv University.     

Understanding tissue specific compositions of bioenergy feedstocks through hyperspectral Raman imaging

Hyperspectral Raman imaging was used to study the tissue/cell type specific distribution of lignin and cellulose polymers within the plant cell walls. Distinct differences in cell wall compositions were identified between two potential bioenergy feedstocks: corn stover and Eucalyptus globulus. Characteristic bands of 627, 1,175, 1,206, and 1,428 cm⁻¹ were only observed for corn stover and 1,381 cm⁻¹ was only present in E. globulus. One‐dimensional and two‐dimensional chemical maps of lignin and cellulose were generated for the stem of corn stover, ranging from the epidermis to the pith area and revealed that lignin and cellulose abundance varies significantly among different cell types in the following order: sclerenchyma cells and tracheids (∼5 times) > epidermal cells (∼3 times) > bundle sheath cells > parenchyma cells. The Raman mapping methods developed on corn stover were also validated on E. globulus and clearly highlighted their difference in lignin composition. Biotechnol. Bioeng. 2011;108: 286–295. © 2010 Wiley Periodicals, Inc.     

Cell wall structure and composition in the comb built by hornets and wasps (Vespinae,hymenoptera)

Morphological, mineralogical, and chemical investigations were undertaken to determine the structure and composition of the cell walls of the comb in the nest of Vespa orientalis, Paravespula germanica, and Vespacrabro. Nests of V. orientalis were from three sites having different soil types, namely, Khamra soil, Gramosol soil, and organically rich soil from the city dump in Tel Aviv. Nests of P. germanica were from areas rich in organic matter, and those of V. crabro, shipped from Austria, were similarly comprised of organic matter. Structure and composition of cell walls in the three species differed; furthermore, grain size in the combs differed from that of particles in the surrounding soil.     

Fractionation of carbohydrates in Arabidopsis root cell walls shows that three radial swelling loci are specifically involved in cellulose production

 Three non-allelic radial swelling mutants (rsw1, rsw2 and rsw3) of Arabidopsisthaliana L. Heynh. were shown to be specifically impaired in cellulose production. Fractionation methods that identify, characterise and quantify some of the major cell wall polysaccharides in small quantities of seedlings demonstrated that changes in the production of cellulose are much more pronounced than changes in the production of non-cellulosic polysaccharides. A crude cell wall pellet was sequentially extracted with chloroform methanol (to recover lipids), dimethyl sulphoxide (starch), ammonium oxalate (pectins) and alkali (hemicelluloses). Crystalline cellulose remained insoluble through subsequent treatments with an acetic/nitric acid mixture and with trifluoroacetic acid. Cetyltrimethylammonium bromide precipitation resolved neutral and acidic polymers in the fractions, and precipitation behaviour, monosaccharide composition and glycosidic linkage patterns identified the major polysaccharides. The deduced composition of the walls of wild-type seedlings and the structure and solubility properties of the major polymers were broadly typical of other dicots. The three temperature-sensitive, radial swelling mutants produced less cellulose in their roots than the wild type when grown at their restrictive temperature (31 °C). There were no significant differences at 21 °C where no radial swelling occurs. The limited changes seen in the monosaccharide compositions, glycosidic linkage patterns and quantities of non-cellulosic polysaccharides support the view that the RSW1, RSW2 and RSW3 genes are specifically involved in cellulose synthesis. Reduced deposition of cellulose was accompanied by increased accumulation of starch. Received: 15 December 1999 / Accepted: 18 January 2000     

Études sur la composition chimique des parois d'une levure candida après croissance sur n paraffine

Résumé La composition chimique des parois d'une levure Candida cultivée sur substrat n paraffine a été examinée. Par rapport à celles de levure du même genre, les parois de cette souche renferment moins d'oses neutres mais davantage de protéines et glucosamine. Une partie importante de cette hexosamine entre dans la constitution de la chitine qui représente 22% de la paroi.Le traitement par l'éthylène diamine permet de séparer trois fractions A, B et C dont les structures apparaissent plus complexes que celles obtenues à partir des parois de levures Saccharomyces et autres Candida.La composition chimique originale de ces parois et les complexes glycoprotéiques particuliers obtenus après différents traitements de la paroi peuvent constituer un caractère d'identification de cette souche.

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Summary The cell wall chemical structure of a Candida yeast strain cultivated on a paraffin substrate has been investigated. By comparison with other yeasts of the same generus, the cell walls of this strain contain less neutral sugar but more proteins and glucosamine. An important part of this aminosugar constitutes chitine which represents 22% of the cell wall.By ethylene diamine treatment three fractions A, B, C are obtained whose structure seems more complex than those obtained from Saccharomyces and other Candida cell walls.The original chemical composition of these cell walls and the particular glycoprotein complexes obtained after different treatments of the cell walls may be used as an identification character of this strain.

     

Characterization of the Sclerotinia sclerotiorum cell wall proteome

We used a proteomic analysis to identify cell wall proteins released from Sclerotinia sclerotiorum hyphal and sclerotial cell walls via a trifluoromethanesulfonic acid (TFMS) digestion. Cell walls from hyphae grown in Vogel's glucose medium (a synthetic medium lacking plant materials), from hyphae grown in potato dextrose broth and from sclerotia produced on potato dextrose agar were used in the analysis. Under the conditions used, TFMS digests the glycosidic linkages in the cell walls to release intact cell wall proteins. The analysis identified 24 glycosylphosphatidylinositol (GPI)‐anchored cell wall proteins and 30 non‐GPI‐anchored cell wall proteins. We found that the cell walls contained an array of cell wall biosynthetic enzymes similar to those found in the cell walls of other fungi. When comparing the proteins in hyphal cell walls grown in potato dextrose broth with those in hyphal cell walls grown in the absence of plant material, it was found that a core group of cell wall biosynthetic proteins and some proteins associated with pathogenicity (secreted cellulases, pectin lyases, glucosidases and proteases) were expressed in both types of hyphae. The hyphae grown in potato dextrose broth contained a number of additional proteins (laccases, oxalate decarboxylase, peroxidase, polysaccharide deacetylase and several proteins unique to Sclerotinia and Botrytis) that might facilitate growth on a plant host. A comparison of the proteins in the sclerotial cell wall with the proteins in the hyphal cell wall demonstrated that sclerotia formation is not marked by a major shift in the composition of cell wall protein. We found that the S. sclerotiorum cell walls contained 11 cell wall proteins that were encoded only in Sclerotinia and Botrytis genomes.